Several clonal strains of rat pituitary tumor cells (GH cells) synthesize and secrete into the medium different amounts of two hormones, prolactin (PRL) and growth hormone (GH). The synthesis of these two proteins can be modulated further by externally added physiological and pharmacological agents. Understanding the mechanism(s) involved in the graded basal level production of the hormone prolactin (PRL) and the regulation of its synthesis by thyrotropin-releasing hormone (TRH), hydrocortisone (HC), estradiol (E2), by the drug 5-bromodeoxyuridine (BrdUrd) and by growth factors such as epidermal growth factor (EGF) in GH cells is the primary objective of this research. Though the model cell system utilized in this investigation is not composed of normal cells, it still responds to phsyiological agents such as TRH, HC, E2 and EGF in the same manner as does the normal pituitary gland. Since distinct cell types are believed to produce specific hormones in the normal pituitary gland, the GH subclones which produce two hormones (PRL and GH) provide a novel model system for the understanding of mechanisms whereby neoplastic cells can express a portion of the genetic information that is kept under strict control in normal cells. The synthesis of PRL by a distinct type of cell in the normal pituitary gland represents a differentiated function. The different GH subclones which produce vastly different amounts of PRL provide an ideal system for the investigation of the cellular mechanisms which lead to the regulated synthesis of such a cell-specific protein and, thus, provide a novel means to understand the process of cellular differentiation. Our results suggest that the graded levels of PRL synthesis by different GH-cell subclones at the basal state and after treatment with TRH or BrdUrd is due to altered transcription of the PRL-specific RNA sequences. The striking effect of BrdUrd on PRL synthesis is being utilized to obtain useful information on the nature of hormone receptor interactions with specific DNA binding sites. The recently observed BrdUrd-induced amplification of the PRL gene will be utilized for the understanding of the mechanism of turning the switch "on" or "off" for the process of gene amplification in this system. As drug-induced amplification seems to be limited within the 20 kb segment within and around the PRL gene-coding sequence, these results suggest that there is/are initiation and termination of such preferential DNA replication site(s) within this region which selectively respond to the drug BrdUrd. Identification of this/ these site(s) will be of significant interest for the understanding of gene amplification in eucaryotic cells.